Fundamental puzzles raised by the immune system include the vast number of expressed variable (V) region sequences, the way in which V sequences become linked to a constant region (C) through a joining region (J) element, how lymphocyte clones switch the expressed CH region, the restriction of gene expression to one allele ("allelic exclusion"), the relation of receptor and secreted immunoglobulin and the basis for the antigen specificity of T cells. To address these problems, we are using molecular cloning techniques to examine the organization of mouse immunoglobulin genes. We recently cloned 16 species of immunoglobulin cDNA, constructed a library of 106 clones spanning the mouse genome, isolated clones bearing three CH genes and over 30 bearing VH genes, determined the structure of the C mu gene, sequenced 2,000 base pairs spanning the JH locus, analyzed VH-JH joining and CH switching in nine plasmacytomas and examined C mu gene expression in B and T lymphoid cell lines. We propose now to analyze the organization of VH genes and part of the CH locus, concentrating on the nature and role of their flanking sequences, to determine a number of VH and V kappa nucleotide sequences, to estimate the total number of V kappa genes by hybridization with 10 cloned V kappa probes, to analyze allelic exclusion by studying lines in which both alleles appear to be expressed to search for the locus of Diversity (DH) genes, which encode most of the third hypervariable regions, to identify the membrane (receptor) segments of C gamma genes and to examine the JH locus and specific VH genes in many T cell lines. These studies should contribute to understanding of the genetic basis for antibody diversity, allelic exclusion and T cell specificity and the mechanism of the recombination events required for immunoglobulin expression.